What is the sequence of bacterial reverse transcriptase and can we clone that gene?
The article at the bottom of the page provides some valuable information on bacterial reverse transcriptase. It seems that the specific sequence of base pairs that code for the RT enzyme differ depending on the bacteria (the two examined in the report are myxobacteria and e.coli). In general, the article provides more information on the "what" than the "how". I'm going to continue to search for details on the processes, but meanwhile, if anyone has questions or additions to make to this page, please feel free.
Here are some of the important points I've gathered from the report:
Important Points from the Report
1. A specific region on the bacterial chromosome codes for portions of the DNA-RNA complex we've been looking at. These portions include the msDNA (the DNA part), msrRNA (the RNA part), and the reverse transcriptase which allows for the creation of this compound. As I said above, the coding sequence for these three key portions differences between what the article refers to as e.coli B and e.coli k12.
2. The G residue (see figure 1 in the article) in the folded RNA "template" serves as the primer for reverse transcriptase instead of the 3' end of the primer as in regular transcription.
3. In order for production of the DNA-RNA compound in e.coli B, the open reading frame has to be intact. The authors also suggest that the sequences that code for the msDNA and msrRNA have to be intact as well.
4. Formation of msDNA is RNA-dependent, which of course, accounts for the use of reverse transcriptase to create it.
5. Finally, and to answer the second part of the above question, the authors claim that the gene coding for reverse transcriptase CAN be cloned. To quote directly:
"It was shown that MoMLV reverse transcriptase entends the msDNA of EC86 from the 3' end to the branch point at the G residue. As with myxobacteria, a single site for Ec86 was located in the chromosome. A...fragment containing this site was cloned in E.coli k12, in which is was expressed" (Lim and Maas, 1989).